The complete mitochondrial genome of Pseudopleuronectes herzensteini

Abstract Pseudopleuronectes herzensteini belonging to Pleuronectiformes (family Pleuronectidae) is important in the fishery industry. However, the molecular biology of this valuable fish has hardly been reported. Thus, here we report the complete mitochondrial genome of P. herzensteini. The mitochondrial DNA (mtDNA) of P. herzensteini is 16,719 bp long and contains 13 mitochondrial protein-coding genes (PCGs), 22 transfer RNA (tRNA) genes, two ribosomal RNA (rRNA) genes, and a putative control region between tRNA-P and tRNA-F distinguished by a single short noncoding region. Phylogenetic analysis using PCGs confirmed that this mtDNA sequence belongs to the family Pleuronectidae. This is the first study reporting the complete mitochondrial genome sequence of P. herzensteini.

Littlemouth flounder (Pseudopleuronectes herzensteini; Jordan & Snyder 1901) is an economically important cold-water fish, belonging to Pleuronectiformes (family Pleuronectidae). P. herzensteini inhabits in East Sea of Korea, Northwest Pacific, Yellow Sea, inland Sea of Japan, and Kuril Islands (FishBase 2021). In South Korea, the annual catch has increased since 1990s (KOSIS 2021). Previous reports on P. herzensteini have mainly focused on ecological characteristics (Takahashi et al. 1995;Han and Kim 1999;Lee et al. 2006;Shimoda et al. 2006;Kobayashi et al. 2015) and the studies on genetic information have scarcely been reported. Thus, here we focused on the complete mitochondrial DNA (mtDNA) sequence of P. herzensteini and identified gene distribution, gene sequences, transfer RNA (tRNA) information, and phylogenetic relationships. This is the first study reporting the complete mtDNA sequence of P. herzensteini including the control regions.
The specimen was provided by the National Institute of Fisheries Science (NIFS, Busan, South Korea) and deposited in the Fisheries Bio-resources storage of NIFS (voucher no. NFRDI-FI-TS-0055174: https://www.nifs.go.kr/frcenter/, Dr. Dong-Gyun Kim, combikola@korea.kr). The gDNA was isolated from fin tissue using the Bead TM Genomic DNA Prep Kit for Animal Tissue (Biofact, Daejeon, South Korea). The cox1 gene was amplified by PCR using the fish universal primer set, FishF2, 5 0 -TCGACTAATCATAAAGATATCGGCAC-3 0 ; FishR2, 5 0 -ACTTCAGGG TG-ACCGAAGAATCAGAA-3 0 (Ward et al. 2005). The amplicons were sequenced by Macrogen (Seoul, South Korea) and the cox1 sequence was compared using the Basic Local Alignment Search Tool (BLAST) of the National Center for Biotechnology Information (NCBI). A BlastN (Johnson et al. 2008) search of the cox1 sequence showed 98.96% similarity. to cox1 sequence (MH032527) of the P. herzensteini, supporting that our flounder is P. herzensteini.
Library preparation for next-generation sequencing (NGS) was performed with MGIEasy DNA library prep kit (MGI, Shenzhen, China). NGS was conducted on the MGISEQ-2000 (MGI) with 150 bp paired-end reads. The raw data were deposited in Sequence Read Archive (SRA) database (SRR18558256). The mitochondrial genome of P. herzensteini was recovered by direct mapping to the P. yokohamae mitogenome (NC_028014) using Geneious ver. 11.1.3 (Kearse et al. 2012). The mtDNA of P. herzensteini showed 16,719 bp long circular DNA, and the sequence has been registered in the GenBank database (ON127848).
Except nad6, other PCGs were transcribed on the positive strand started with an ATG codon (nad1, nad2, cox2, atp8, atp6, cox3, nad3, nad4l, nad4, nad5, nad6, and cob) except cox1 which started with a GTG codon. The nad2, cox2, nad3, nad4, and cob were terminated with the truncated codons T-. The remaining six PCGs were stopped with TAA codon except for nad5 and nad6 where TAG codon was present. The results showed 22 tRNA genes including two tRNA-L and two tRNA-S. Except for seven tRNA genes (tRNA-A, tRNA-N, tRNA-C, tRNA-Y, tRNA-S2, tRNA-E, and tRNA-P), other 15 tRNA genes were encoded on the positive strand. The D-arm loop structure in the tRNA-C and tRNA-S1 was not observed; however, the rest tRNAs retained the standard cloverleaf structure in the predicted secondary structure.
Small rRNA with a 949 bp length was located in between tRNA-F and tRNA-V, whereas large rRNA was located in between tRNA-V and tRNA-L2 and had a length of 1716 bp. The putative control region was 1016 bp long and located in between tRNA-P and tRNA-F. The gene order was accorded with P. yokohamae (Zheng et al. 2016).
Several nucleotide sequences of PCGs from other related species, divided by family level for comparison with P. herzensteini, were collected from NCBI and Acipenseridae was utilized as an out-group. The phylogenetic tree was constructed using the MEGA 11 software (Tamura et al. 2021). The P. herzensteini was classified with P. yokohamae belonging to Pleuronectidae (Figure 1). Furthermore, P. herzensteini was gathered with Pleuronectidae not Paralichthyidae and Bothidae. The results of the present study evidence that the flounder analyzed in this study belongs to P. herzensteini.

Data availability statement
The genome sequence data that support the findings of this study are openly available in GenBank of NCBI at https://www.ncbi.nlm.nih.gov/ under the accession no. ON127848. The associated BioProject, SRA, and Bio-Sample numbers are PRJNA821835, SRR18558256, and SAMN27124384, respectively.